Transcobalamin II deficiency in twins with a novel variant in the TCN2 gene: case report and review of literature

Case 1

A 4-month-old (corrected age: 2 months) female was admitted to our hospital with fever, respiratory distress, vomiting, diarrhea, and failure to thrive.

In medical history, birth weight, length, and head circumference were 1445 g (10–25th percentile) and 42 cm (10–25th percentile), respectively. She had been followed up in a newborn intensive care unit because of prematurity for 50 days. She was discharged from intensive care unit with 2.8 kg (10–25th percentile) weight and 47 cm (10th percentile) length.

In physical examination, pallor, tachycardia, and growth retardation (weight: 3.8 kg, <3 percentile; length: 53 cm, 3–10th percentile) were detected. The patient was hypotonic with no head control. No dysmorphic appearance noted. The other system examination findings were all normal.

In laboratory investigation, bicytopenia was revealed (white blood cell count, 6.3 × 10⁹/L; absolute neutrophil count, 0.85 × 10⁹/L; hemoglobin level, 7.2 g/dL; platelet count, 156 × 10⁹/L). Mean corpuscular volume (97.1 fL) showed macrocytic anemia. C-reactive protein level was 14 mg/mL (range, 0–5 mg/mL). Lymphocyte subsets were within the normal range. Low IgG (33.1 mg/dL), IgA (<27.8 mg/dL), and IgM (19.4 mg/dL) levels were noted (Table 1). Serum electrolyte levels, renal and liver function tests were within the normal range. Urine and stool analyses were normal. Bone marrow aspiration was performed to evaluate the hematological findings. Bone marrow smear revealed megaloblastic changes which include giant pronormoblasts, nucleocytoplasmic dissociation, giant myeloid cells, binucleated normoblasts, nucleocytoplasmic dissociation in normoblasts (Figure 1B–D). Owing to hypotonia and development delay, brain magnetic resonance imaging (MRI) was performed and pattern of retarded myelination in white matter due to prematurity was revealed.

In the clinical follow-up, antimicrobial therapy initiated for pneumonia. Owing to anemia and low IgG levels, red blood cell transfusion and intravenous immunoglobulin (IVIG) infusion were administrated. The patient started feeding with an amino acid–based formula. However, vomiting, diarrhea, anemia, and neutropenia persisted.

Further biochemical investigations revealed elevated serum homocysteine levels (44.8 mmol/L [5], [6], [7], [8], [9], [10], [11], [12], [13], [14], [15]) and elevated urine methylmalonic acid levels (756.4 μmol/L [<3.6]). Plasma ammonia and lactate levels were within the normal range. Serum vitamin B12 and folate levels were 777 pmol/L (147–664) and 16.6 nmol/L (11.3–47.6), respectively (Table 1). With all these findings, the inborn error of cobalamin metabolism was suspected, and hydroxy-Cbl (1 mg intramuscular [i.m.] once daily) initiated.

After 1-week 1 mg i.m. once daily hydroxy-Cbl treatment, hematologic parameters and homocysteine levels (5.1 mmol/L) normalized. Urine methylmalonic acid level decreased to 2.1 μmol/L. Immunoglobulin levels were improved (Table 1). No transfusion required. On the last admission to the outpatient clinic, at nine months old (corrected age: 7 months), neurological examination findings were normal. In Denver developmental screening test II, personal-social, fine motor–adaptive, language and gross motor skills were compatible with 6, 7, 7, and 8 months age, respectively. Growth parameters were at the 10–25th percentile, and the hematological parameters were within the normal range.

Case 2

A 4-month-old (corrected age: 2 months) male infant was admitted to our hospital with vomiting, diarrhea, and failure to thrive.

In medical history, birth weight, length, and head circumference were 1200 g (<3rd percentile) and 39 cm (3–10th percentile), respectively. He had been followed up in a newborn intensive care unit because of prematurity for 50 days. He was discharged from intensive care unit with 2800 g (3–10th percentile) weight and 47 cm (3–10th percentile) length.

The physical examination findings were pallor, tachycardia, and growth retardation (weight: 3.6 kg, <3rd percentile; length: 54 cm (3–10th percentile)). The patient was hypotonic with no head control, and feeding difficulty was present. No dysmorphic appearance noted. The other system examination findings were all normal.

In laboratory evaluation, pancytopenia was revealed (white blood cell count, 4.8 × 10⁹/L; absolute neutrophil count, 0.58 × 10⁹/L; hemoglobin level, 7.0 g/dL; platelet count, 12 × 10⁹/L). Mean corpuscular volume (94 fL) showed macrocytic anemia. Low IgG (32.9 mg/dL), IgA (<27.8 mg/dL), and IgM (<17.3 mg/dL) levels were detected. Lymphocyte subsets were unremarkable (Table 1). Serum electrolyte levels and liver and kidney function test results were within the normal range. Urine and stool analyses were normal. Giant pronormoblasts, nucleocytoplasmic dissociation, giant myeloid cells, binucleated normoblasts, nucleocytoplasmic dissociation in normoblasts were detected in bone marrow smear (Figure 1B–D). These findings were compatible with megaloblastic changes. Brain MRI showed pattern of retarded myelination in white matter due to prematurity.

In the clinical follow-up, red blood cell and platelet transfusions, and IVIG infusion were repeatedly administrated. However, no hematological improvement observed. Despite feeding with an amino acid–based formula, vomiting, diarrhea, anemia, neutropenia, and thrombocytopenia persisted.

Further biochemical investigations revealed elevated serum homocysteine levels (42.6 mmol/L [<15]) and elevated urine methylmalonic acid levels (843.1 μmol/L). Plasma ammonia and lactate levels were within the normal range. Serum vitamin B12 and folate levels were 446 pmol/L (147–664) and 15.7 nmol/L (11.3–47.6), respectively (Table 1). Hydroxy-Cbl therapy (1 mg, i.m., once daily) initiated.

Hematologic parameters and homocysteine levels (5.5 mmol/L) normalized with the 1 week treatment of 1 mg i.m. once daily hydroxy-Cbl. Vomiting and diarrhea regressed, and immunoglobulin levels improved. Urine methylmalonic acid level decreased to 3.2 μmol/L (Table 1). In the clinical follow-up, no transfusions required. On the last admission to the outpatient clinic, at nine months old (corrected age: 7 months), neurological examination revealed normal findings. In Denver developmental screening test II, personal-social, fine motor–adaptive, language and gross motor skills were compatible with 7, 6, 6, and 7 months age, respectively. Growth parameters were at the 10–25th percentile, and hematological parameters were within the normal range with weekly i.m. hydroxy-Cbl treatment.

Genetic analysis

TCN2 gene mutation analysis was performed by sequencing of the coding exons and the exon-intron boundaries of the genes. Genomic DNA was isolated from peripheral blood cells with QIAGEN DNA Blood Mini Kit in accordance with the protocol provided with the kit. Sequencing was performed with MiSeq V2 chemistry on MiSeq instrument (Illumina, CA, USA). A novel mutation presented here was predicted to be disease causing by in silico analysis NextGENe software (SoftGenetics, State College, PA, USA).

In both cases, novel pathogenic homozygous c.241C>T (p.Gln81Ter) variant in exon two of the TCN2 gene was revealed (Figure 1D). These mutations are predicted to be disease causing by in silico analysis software. Therefore, this variant might possibly be a likely pathogenic variant. The variant has not been reported in any public database (Genome Aggregation Database and Exome Aggregation Consortium Database).